PILOT-SCALE PURIFICATION OF HUMAN MONOCLONAL IGM (COU-1) FOR CLINICAL-TRIALS

No standard procedure is available for the purification of human monoc lonal antibodies for human i.v. administration, Here we describe the p rocedure developed for pilot scale purification of the human IgM monoc lonal antibody COU-1 directed against a cancer-associated antigen. The hybridoma cells were grown in protein-free medium and purification fr om the clarified culture supernatant was carried out in 4 simple chrom atographic steps: (1) hydroxylapatite chromatography; (2) hydrophobic interaction chromatography on phenyl-Sepharose: (3) cation-exchange ch romatography on sulphonyl-Sepharose; and (4) anion-exchange chromatogr aphy on tetraethylamino-Sepharose. The product was substantially pure with regard to protein after step 3, but contained DNA which was remov ed in step 4. The average recovery of the IgM was 54% with a range of 40-65%. Importantly, the ability of the antibody to bind to its antige n in ELISA was fully maintained during the purification. Subsequently, the purified antibody was isotope labelled and successfully used for in vivo detection of colon, rectal and pancreas carcinomas in patients . The purification procedure described appears to compare favourably w ith previously published methods, but a critical comparison is not pos sible due to the lack of necessary information in the available litera ture.